Phenotypic screening of seed retention and histological analysis of the abscission zone in Festuca pratensis and Lolium perenne

Background Seed retention is the basic prerequisite for seed harvest. However, only little breeding progress has been achieved for this trait in the major forage grasses. The aim of this study was to evaluate the potential of plant genetic resources of the important fodder grasses Festuca pratensis Huds. and Lolium perenne L. as source for seed retention in the breeding process. Furthermore, the morphology of the abscission zone, where shattering occurs, was studied on the cell tissue level in different developmental stages of contrasting accessions. Results 150 and 286 accessions of Festuca pratensis and Lolium perenne were screened for seed retention, respectively. Contrasting accessions were selected to be tested in a second year. We found a great variation in seed retention in Festuca pratensis and Lolium perenne, ranging from 13 to 71% (average: 35%) and 12 to 94% (average: 49%), respectively, in the first year. Seed retention was generally lower in the second year. Cultivars were within the accessions with highest seed retention in Festuca pratensis, but had lower seed retention than ecotypes in Lolium perenne. Field-shattered seeds had a lower thousand grain weight than retained seeds. Cell layers of the abscission zone appeared already in early seed stages and were nested within each other in accessions with high seed retention, while there were two to three superimposed layers in accessions with low seed retention. Conclusions Plant genetic resources of Lolium perenne might be a valuable source for breeding varieties with high seed retention. However, simultaneous selection for high seed weight is necessary for developing successful commercial cultivars. Supplementary Information The online version contains supplementary material available at 10.1186/s12870-024-05231-0.


Supplements
Table S1: Protocol for combined conventional and microwave-proceeded fixation, dehydration and resin embedding of the transition area from the rachis to the developing caryopsis for histological analysis.Microwave processing (steps 1-3) in a PELCO Bio Wave®34700-230 (Ted Pella, Inc., Redding CA, USA).

Process
Reagent Power [W]

Climate data Malchow
Climate data (temperature, precipitation, wind velocity and maximum speed) were obtained from the IPK weather station, located about 3 km south-east of the trial location.

Climate data Steinach
Climate data (temperature, precipitation and wind velocity) were obtained from the weather station of the Bayerische Landesanstalt für Landwirtschaft (LfL), located about 2 km north of the trial location.

Trial with Lolium multiflorum var. westerwoldicum
In L. multiflorum (diploid), the seed retention of families from two different selection processes, for high seed retention (V3) and for low seed retention (V4), was tested.Seeds from groups of plants with similar seed retention were grown and polycrossed.Progenies were tested again for seed retention and used as parents for V3 and V4 if seed retention was > 70 % or < 30 %, respectively.The first plants originate from a chemical mutagenesis experiment in order to select for genotypes with higher seed retention.L. multiflorum was used due to the lack of need of a vernalisation period and therefore faster breeding process.
Plants were sown in 2017 in Bocksee in the north of Germany.
On average, seed retention was 76 % in V3 and 60 % in V4 before agitation.Agitation after harvest resulted in the detachment of 15 % and 22 % of the still attached seeds in V3 and V4, respectively.Variability between families was high with average seed retentions in the field between 63 % and 83 % in V3, and 45 % and 73 % in V4.V3 (A) was selected for high seed retention, V4 (B) was selected for low seed retention.
Boxes represent first and third quartile with a line for the median.Whiskers extend to minimum and maximum values within the 1.5 times interquartile range.Dots display outliers.
Thousand grain weight was determined for some members of the Lolium multiflorum families V3 and V4.TGW was on average higher in the families selected for low seed retention (V4) (p = 0.005; Fig. S5).The field-shattered seeds had a lower TGW than the hand-stripped seeds with seeds shattered by agitation ranging between both groups in V3 and in V4 (p < 0.001).

Fig. S1 :
Fig. S1: Wind velocity (mean and maximum) per day, sum of monthly precipitation and temperature mean per month during the field experiment in Malchow.Period of heading (HAE) and harvest are given.

Fig. S2 :
Fig. S2: Wind velocity (mean) per day, sum of monthly precipitation and temperature mean per month during the field experiments in Steinach.

Fig. S3 :
Fig. S3: Regressions between seed retentions in 2018 and 2019 in Malchow for Festuca pratensis and Lolium perenne in the groups selected for 2019 with high, medium and low seed retention.

Fig. S5 :
Fig. S5: Thousand grain weight of seeds from field-shattered, shattered by agitation and handstripped fractions of Lolium multiflorum families (n=53 (V3: selected for high seed retention) and n=50 (V4: selected for low seed retention)).Boxes represent first and third quartile with a line for the median.Whiskers extend to minimum and maximum values within the 1.5 times interquartile range.Dots display outliers.

Fig. S7 :
Fig. S7: Histological characterisation of the transition area from the rachilla to the developing caryopsis of Festuca pratensis at different developmental stages.Light microscopy images of 17EP1166-37/8 (A-D) and Cosmopolitan (E-H) with high seed retention and 17EP1166-33/5 (I-L) and GR 1835 (M-P) with low seed retention (I-P).Red box indicates the abscission zone and arrowheads indicate a fracture surface on the rachilla.Ca, caryopsis; Ra, rachilla; Vc, vasculature.

Table S2 :
Protocol for sample preparation of the transition area from the rachis to the

each step of dehydration additional 15 min on shaker 6. Critical Point Drying
of samples in a BAL-TEC CPD 030